Film-forming substances are widely used in compositions for skin care and hair care as conditioning agents and moisturizers, and to protect the skin and hair against environmental and chemical damage. These substances adsorb onto and/or absorb into the skin or hair, forming a protective coating. Commonly used film-forming substances include synthetic polymers, such as silicones, polyvinylpyrrolidone, acrylic acid polymers, and polysaccharides, and proteins, such as collagen, keratin, elastin, and silk proteins. Proteins are known to be particularly effective film-forming agents. Because of their low solubility at the conditions used in skin and hair care products, proteins are commonly used in the form of peptides, formed by the hydrolysis of the protein.
In hair care compositions, film-forming substances are used to form a protective film on the surface of the hair to protect it from damage due to grooming and styling, shampooing, and exposure to ultraviolet light and the reactive chemicals commonly used in permanent wave agents, hair coloring products, bleaches, and hair straighteners, which denature the hair keratin protein. Moreover, these film-forming substances improve the elasticity of the hair. Film-forming substances that have been used in hair care products include proteins, such as keratin and collagen and hydrolysates thereof, and polymeric materials, such as polyacrylates, long chain alkyl quaternized amines, and siloxane polymers. For example, Cannell at al. in U.S. Pat. No. 6,013,250 describe a hair care composition for treating hair against chemical and ultraviolet light damage. This composition comprises hydrolyzed protein, having an abundance of sulfur containing amino acids, and divalent cations. Omura et al. in U.S. Pat. No. 6,139,851 describe a hair care cosmetic for treating split ends which contains one or more types of silicone derivatives, one or more types of specific polyether modified silicone, and a lower alcohol. The major problem with these compositions is that they lack the required durability required for long-lasting protection.
Film-forming substances are also used in skin care compositions to form a protective film on the skin. This film lubricates and coats the skin to passively impede the evaporation of moisture and smoothes and softens the skin. Commonly used film-forming substances in skin care compositions include hydrolyzed animal and vegetable proteins (Puchalski et al., U.S. Pat. No. 4,416,873, El-Menshawy et al., U.S. Pat. No. 4,482,537, and Kojima et al., JP 02311412).
Silk proteins have also been used as film-forming substances in skin care and hair care compositions. Because of the low solubility of these proteins, they are used in the form of protein hydrolysates. Silk proteins are ideally suited for film-forming and coating applications because of their ability to self-assemble in solution (Winkler et al., Int. J. Biol. Macromol. 24:265–270 (1999)). This self-assembly property of silk proteins is due to the formation of anti-parallel beta-pleated sheets via hydrogen bonding and hydrophobic interactions (Arcidiacono et al., Appl. Microbiol. Biotechnol. 49:31–38 (1998)). Silk proteins are produced by over 30,000 species of spiders and by many insects particularly in the order Lepidoptera (Foelix, R. F. Biology of Spiders, Harvard University Press Cambridge, Mass. (1992)). Few of these silk proteins have been studied in detail. The cocoon silk of the domesticated silkworm Bombyx mori, i.e. silk fibroin, and the dragline silk of the orb-weaving spider Nephila clavipes are among the best characterized.
Recombinant DNA technology has been used to provide a more practical source of silk proteins. Ohshima et al. (Proc. Natl. Acad. Sci. USA, 74:5363–5367 (1977)) report the cloning of the silk fibroin gene complete with flanking sequences of the silkworm Bombyx mori into E. coli Petty-Saphon et al. (EP 0230702) disclose the recombinant production of silk fibroin and silk sericin from a variety of hosts including E. coli, Sacchromyces cerevisiae, Pseudomonas sp., Rhodopseudomonas sp., Bacillus sp., and Strepomyces sp. The production of recombinant spider silk proteins is also known. Xu et al. (Proc. Natl. Acad. Sci. U.S.A., 87:7120–7124 (1990)) report the determination of the sequence for a portion of the repetitive sequence of a dragline spider silk protein, Spidroin 1, from the spider Nephila clavipes, based on a partial cDNA clone. Hinman and Lewis (J. Biol. Chem. 267:19320–19324 (1992)) report the sequence of a partial cDNA clone encoding a portion of the repeating sequence of a second fibroin protein, Spidroin 2, from dragline silk of Nephila clavipes. Lewis et al. (U.S. Pat. Nos. 5,728,810 and 5,989,894) disclose the expression of spider silk proteins including protein fragments and variants of Nephila clavipes from transformed E. coli. cDNA clones encoding minor ampullate spider silk proteins and the expression thereof is described by Lewis et al. (U.S. Pat. Nos. 5,733,771 and 5,756,677). Lewis et al. (U.S. Pat. No. 5,994,099) describe the cloning of cDNA encoding the flagelliform silk protein from an orb-web spinning spider. Fahnestock (U.S. Pat. No. 6,268,169) describes novel spider silk analog proteins derived from the amino acid consensus sequence of repeating units found in the natural spider dragline of Nephila clavipes. The synthetic spider dragline was produced from E. coli, Bacillus subtilis, and Pichia pastoris recombinant expression systems. Lewis et al. (WO 03/020916) describe the cloning of spider silk proteins from the major ampullate glands of Nephila madagascariensis, Nephila senegalensis, Tetragnatha kauaiensis, Tetragnatha versicolor, Argiope aurantia, Argiope trifasciata, Gasteracantha mammosa, and Latrodectus geometricus, the flagelliform glands of Argiope trifasciata, the ampullate glands of Dolomedes tenebrosus, two sets of silk glands from Plectreurys tristis, and the silk glands of the mygalomorph Euagrus chisoseus. 
The solubility of recombinant silk proteins in aqueous solution depends on the type of silk protein as well as the expression system used. Recombinant silkworm silk proteins are expressed in insoluble form and can only be dissolved using harsh solvents. Recombinant spider silk proteins are expressed in soluble form in bacterial hosts and in both soluble and insoluble forms in yeasts. However, the recombinant spider silk proteins that are expressed in soluble form in microbial systems become insoluble upon purification and are extremely difficult to resolubilize after drying or precipitation (Arcidiacono et al., Macromolecules 35:1262–1266 (2002)), limiting applications which require processing of the proteins into different types of fibers, films or coatings. Winkler et al. (Int. J. Biol. Macromol. 24:265–270 (1999)) report that recombinant spider silk proteins rapidly self-assemble upon purification to form insoluble microfibrils. Fahnestock (Rev. Mol. Biotechnol. 74:105–119 (2000)) also reports that once spider silk analog proteins were precipitated, they could only be redissolved in denaturing solvents, such as aqueous quanidine hydrochloride or hexafluoroisopropanol. However, a purification method which results in a silk protein or analog protein that can be redissolved in water after precipitation is described by Fahnestock et al. (copending, co-owned U.S. patent application Ser. No. 10/704,337, filed Nov. 7, 2003, entitled “A Method for Purifying and Recovering Silk Proteins in Soluble Form and Uses Thereof”). In that method, the silk protein is precipitated from the cell extract by the addition of a salt, such as ammonium sulfate, at a temperature below 20° C. The recovered protein is readily redissolved in water.
The water solubility of spider silk proteins expressed in plants is reported to be much higher than that of the microbially expressed proteins, as they are readily soluble in aqueous buffers (Scheller et al. DE 10113781). Additionally, Lazaris et al. (Science 295:472–476 (2002)) describe the production of soluble recombinant spider silk protein produced in mammalian cells. After precipitation, this spider silk protein was readily redissolved in phosphate-buffered saline. The solubility of the spider silk proteins produced in mammalian cells was attributed to the presence of the COOH-terminus in these proteins, which makes them more hydrophilic. These COOH-terminal amino acids are absent in spider silk proteins expressed in microbial hosts.
There has been no description in the art of the use of soluble, intact silk proteins in skin care, hair care, or hair coloring compositions. Silk proteins from the silkworm Bombyx mori have been used in cosmetic compositions including hair care products. However, these silk proteins are only soluble in harsh solvents that are not compatible with skin care and hair care products. Consequently, these silk proteins have been used in the form of silk protein peptides, formed by the hydrolysis of the silk protein (Kuroda et al., JP 309816; Terada et al., JP 08027186; Otoi et al., JP 63092671; Inoe et al., JP 2574732; and Yamaguchi et al., JP 07067687). Morelle et al. in U.S. Pat. No. 5,504,228 describe the use of acylated silk fibroin hydrolysates in cosmetics. Additionally, Oshika et al. in U.S. Pat. No. 5,747,015 describe a hair care product which comprises a salt of an acylated compound obtained by condensing fatty acids with silk protein-derived peptides, which were obtained by the hydrolysis of silk protein. While providing some beneficial coating effect, the silk protein peptides are not as effective as film-forming agents as the soluble, intact proteins. Ritter et al. (DE 3139438) describe the use of colloidal silk protein as an additive in hair care products. However, the colloidal silk protein is not as effective in film forming and coating for skin or hair treatment as a soluble, intact silk protein.
Philippe et al. in U.S. Pat. No. 6,280,747 describe the use of natural or recombinant spider silk proteins in cosmetic and dermatological compositions such as hair care, skin care, make-up, and sunscreen products. However, the spider silk proteins described in that disclosure are not water-soluble. Therefore, the beneficial effects of the self-assembly and coating properties of the spider silk proteins are not fully realized.
In view of the above, there is a need for a novel class of hair care and hair coloring compositions that provide superior durability and long-lasting protection from the various activities mentioned above that cause hair damage. Additionally, there is a need for a novel class of skin care compositions that provide a superior smooth and durable, protective coating as well as added strength to the skin.